Antitumor Activity of Antimicrobial Peptides Containing CisoDGRC in CD13 Negative Breast Cancer Cells

Lei Hou,Min Meng,Xinhan Zhao,Caigang Liu,Pei Wang,Qian Ning

doi:10.1371/journal.pone.0053491

Lei Hou, Min Meng + Show 4 more

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https://doi.org/10.1371/journal.pone.0053491

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Backgroud isoAsp-Gly-Arg (isoDGR) is a derivative of the Asn-Gly-Arg (NGR) motif, which is used as a targeted delivery tool to aminopeptidase N (CD13) positive cells. Recent studies have shown that cyclic isoDGR (CisoDGRC) has a more efficient affinity with αvβ3, a type of integrin that overexpresses in tumor cells. Antimicrobial peptides (AMPs) are an efficient antitumor peptide that specifically kills tumor cells. In the present study, we designed antimicrobial peptides containing the CisoDGRC motif (CDAK) and assessed its antitumor activity for CD13−/αvβ3 + breast cancer cells (MCF-7 and MDA-MB-231) in vitro and in vivo.MethodsIn vitro: We assessed the cytotoxicity of CDAK for MCF-7 and MDA-MB-231 breast cancer cells, the human umbilical vein endothelial cell (HUVEC), and human foreskin fibroblasts (HFF). We performed an apoptosis assay using Annexin-V/PI, DNA ladder, mitochondrial membrane potential, and Caspase-3 and Bcl-2. The effect on cell cycles and affinity with cell were tested using flow cytometry and fluorescent microscopy and the effect on invasion was analyzed using an invasion assay. CDAK was injected intravenously into tumor-bearing athymic nude mice in vivo experiment.ResultsCDAK showed cytotoxic activity in MCF-7 and MDA-MB-231 cells, whereas HUVEC and HFF were less sensitive to the peptides. CDAK induced apoptosis, reduced mitochondrial membrane potential, promoted Caspase-3, and inhibited Bcl-2 expression in the two breast cancer cell lines. In addition, CDAK inhibited proliferation of cancer cell through S phase arrest, and own selective affinity with MCF-7 and MDA-MB-231cells, inhibited the invasion of MDA-MB-231 cells. In vivo, CDAK significant inhibited the progression of the tumor and the generation of neovascularization.ConclusionAntimicrobial peptides containing the CisoDGRC (CDAK) motif could efficiently exhibit the antitumor activity for CD13−/αvβ3 + breast cancer cells.

Highlights

Various compounds and particles have been coupled or added synthetically to the Asn-Gly-Arg (NGR) motif to target recognized aminopeptidase N (CD13), which are over-expessed in angiogenic blood vessels and some tumors cells, which increases antitumor activity
CDAK showed cytotoxic activity in MCF-7 and MDA-MB-231 cells, whereas human umbilical vein endothelial cell (HUVEC) and human foreskin fibroblasts (HFF) were less sensitive to the peptides
We firstly examined the expression of CD13 and avb3 on the MCF-7, MDA-MB-231, HUVEC, and Fibroblast cells using Western-blot

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Various compounds and particles have been coupled or added synthetically to the Asn-Gly-Arg (NGR) motif to target recognized aminopeptidase N (CD13), which are over-expessed in angiogenic blood vessels and some tumors cells, which increases antitumor activity. This includes chemotherapeutic drugs, gene molecular, and pro-apoptosis peptides [1,2,3]. Studies have shown that the cyclic isoDGR (CisoDGRC) motif with a disulfide bridge constraint can increase the tumor targeting efficiency and stability of bent conformation [13]. CisoDGRC appears to be a promising candidate as a tool for targeted delivery of anti-agents for CD132/avb3+ tumor cells, such as MCF-7 and MDA-MB-231 cell lines [14,15]

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